Long-term tolerance to factor VIII is achieved by administration of interleukin-2/interleukin-2 monoclonal antibody complexes and low dosages of factor VIII

Summary: Background: Regulatory T cells (Tregs) play a pivotal role in regulating anti-factor VIII (FVIII) immune responses. Interleukin (IL)-2 mixed with a particular IL-2 monoclonal antibody (mAb; JES6-1) can induce the selective expansion of Tregs in vivo. Methods: In the prevention experiments, we treated mice with hemophilia A with IL-2/IL-2mAb complexes (three times per week) and concurrently with FVIII protein (80 U kg^-1 per week) for 4 weeks. Generation of anti-FVIII immune responses was examined afterward. Next, to induce long-term tolerance to FVIII, a series of treatment dosages and schedules for administering IL-2/IL-2mAb complexes and FVIII protein in mice with hemophilia A was evaluated. Results: Compared to control mice that were treated with only FVIII, which produced high-titer anti-FVIII antibodies, mice treated with IL-2/IL-2mAb complexes plus FVIII produced no antibodies. A marked seven-fold increase in CD4⁺CD25⁺Foxp3⁺Helios⁺ natural Tregs was maintained for 4 weeks in blood, spleen, and lymph nodes and then dropped to normal levels within the next 10 days. The suppressive functions of expanded Tregs were demonstrated with suppressive, proliferative, and cytokine assays. The administration of anti-CD25 mAb (PC-61) blocked this protective effect, confirming the involvement of Tregs in suppressing anti-FVIII immune responses. Importantly, administration of IL-2/IL-2mAb complexes (three times per week for 8 weeks) combined with contiguous weekly injections of low-dosage FVIII protein (20 U kg^-1 per week for 18 weeks) not only abrogated the formation of anti-FVIII antibodies but also induced long-term tolerance to FVIII. Conclusions: Treatment with IL-2/IL-2mAb complexes is highly promising for the induction and maintenance of FVIII-specific tolerance after FVIII protein replacement therapy.